phosphorus-radioisotopes and Obesity

Topics: Adenosine Triphosphate; Adipose Tissue; Animals; Brain; Calcium; Carnitine; DNA; Enzyme Activation; Fatty Acids, Nonesterified; In Vitro Techniques; Insulin; Kidney; Liver; Magnesium; Mice; Mice, Inbred Strains; Mitochondria, Liver; Myocardium; Neurospora crassa; Obesity; Oleic Acids; Palmitic Acids; Phosphorus Radioisotopes; Pyruvate Dehydrogenase Complex; Pyruvates; Rats; Swine

Protein tyrosine phosphatases (PTPs) are required for the dephosphorylation of the insulin receptor (IR) and its initial cellular substrates, and it has recently been reported that PTP-1B may play a role in the pathogenesis of insulin resistance in obesity and type 2 diabetes mellitus (DM). We therefore determined the amount and activity of PTP-1B in abdominal adipose tissue obtained from lean nondiabetic subjects (lean control (LC)), obese nondiabetic subjects (obese control (OC)), and subjects with both type 2 DM (DM2) and obesity (obese diabetic (OD)). PTP-1B protein levels were 3-fold higher in OC than in LC (1444 +/- 195 U vs 500 +/- 146 U (mean +/- SEM), P < .015), while OD exhibited a 5.5-fold increase (2728 +/- 286 U, P < .01). PTP activity was assayed by measuring the dephosphorylating activity toward a phosphorus 32-labeled synthetic dodecapeptide. In contrast to the increased PTP-1B protein levels, PTP-1B activity per unit of PTP-1B protein was markedly reduced, by 71% and 88% in OC and OD, respectively. Non-PTP-1B tyrosine phosphatase activity was comparable in all three groups. Similar results were obtained when PTP-1B activity was measured against intact human IR. A significant correlation was found between body mass index (BMI) and PTP-1B level (r = 0.672, P < .02), whereas BMI and PTP-1B activity per unit of PTP-1B showed a strong inverse correlation (r = -0.801, P < .002). These data suggest that the insulin resistance of obesity and DM2 is characterized by the increased expression of a catalytically impaired PTP-1B in adipose tissue and that impaired PTP-1B activity may be pathogenic for insulin resistance in these conditions.

Topics: Adipose Tissue; Adult; Aged; Animals; Blotting, Western; Cell Line; Diabetes Mellitus, Type 2; Enzyme Activation; Female; Fibroblasts; Humans; Hydrolysis; Male; Middle Aged; Nerve Tissue Proteins; Obesity; Phosphorus Radioisotopes; Phosphorylation; Precipitin Tests; Protein Tyrosine Phosphatases; Rats; Receptor-Like Protein Tyrosine Phosphatases, Class 5

Hepatocyte membranes from both lean and obese Zucker rats exhibited adenylate cyclase activity that could be stimulated by glucagon, forskolin, NaF and elevated concentrations of p[NH]ppG. In membranes from lean animals, functional Gi was detected by the ability of low concentrations of p[NH]ppG to inhibit forskolin-activated adenylate cyclase. This activity was abolished by treatment of hepatocytes with either pertussis toxin or the phorbol ester TPA, prior to making membranes for assay of adenylate cyclase activity. In hepatocyte membranes from obese animals no functional Gi activity was detected. Quantitative immunoblotting, using an antibody able to detect the alpha subunit of Gi, showed that hepatocyte plasma membranes from both lean and obese Zucker rats had similar amounts of Gi-alpha subunit. This was 6.2 pmol/mg plasma membrane for lean and 6.5 pmol/mg plasma membrane for obese animals. Using thiol pre-activated pertussis toxin and [32P]-NAD+, similar degrees of labelling of the 40 kDa alpha subunit of Gi were found using plasma membranes of both lean and obese Zucker rats. We suggest that liver plasma membranes from obese Zucker rats express an inactive Gi alpha subunit. Thus lesions in liver Gi functioning are seen in insulin-resistant obese rats and in alloxan- and streptozotocin-induced diabetic rats which also show resistance as regards the acute actions of insulin. Liver plasma membranes of obese animals also showed an impairment in the coupling of glucagon receptors to Gs-controlled adenylate cyclase, with the Kd values for activation by glucagon being 17.3 and 126 nM for lean and obese animals respectively. Membranes from obese animals also showed a reduced ability for high concentration of p[NH]ppG to activate adenylate cyclase. The use of [32P]-NAD+ and thiol-preactivated cholera toxin to label the 43 kDa and 52 kDa forms of the alpha-subunit of Gs showed that a reduced labelling occurred using liver plasma membranes from obese animals. It is suggested that abnormalities in the levels of expression of primarily the 52 kDa form of alpha-Gs may give rise to the abnormal coupling between glucagon receptors and adenylate cyclase in liver membranes from obese (fa/fa) Zucker rats.

Topics: Adenosine Diphosphate Ribose; Adenylate Cyclase Toxin; Adenylyl Cyclases; Animals; Blotting, Western; Cell Membrane; Cholera Toxin; Colforsin; Diabetes Mellitus, Experimental; Glucagon; GTP-Binding Proteins; Guanylyl Imidodiphosphate; Insulin Resistance; Liver; Male; NAD; Obesity; Pertussis Toxin; Phosphorus Radioisotopes; Rats; Rats, Inbred Strains; Rats, Zucker; Receptors, Gastrointestinal Hormone; Receptors, Glucagon; Virulence Factors, Bordetella